Surface-enhanced raman scattering biosensor

ABSTRACT

A general purpose sensor architecture integrating a surface enhanced Raman spectroscopy (SERS) substrate, a diffractive laser beam delivery substrate and a diffractive infrared detection substrate is provided that can be used to implement a low-cost, compact lab-on-a-chip biosensor that can meet the needs of large-scale infectious disease testing. The sensor architecture can also be used in any other application in which molecules present in the liquid, gaseous or solid phases need to be characterized reliably, cost-effectively and with minimal intervention by highly skilled personnel.

BACKGROUND OF THE INVENTION

This application is a continuation of U.S. patent application Ser. No.17/500,700, filed on Oct. 13, 2021, which claims priority to thefollowing U.S. Provisional Applications: Ser. No. 63/090,917, filed Oct.13, 2020; Ser. No. 63/092,212, filed Oct. 15, 2020; Ser. No. 63/112,672,filed Nov. 12, 2020; Ser. No. 63/112,674, filed Nov. 12, 2020; Ser. No.63/126,075, filed Dec. 16, 2020; and Ser. No. 63/151,650, filed Feb. 20,2021. The entire disclosures of the aforementioned applications areincorporated herein by reference.

FIELD OF THE INVENTION

The present invention relates generally to optical sensors and, moreparticularly, to a holographic waveguide sensor for molecular detection.

BACKGROUND OF THE RELATED ART

There is a growing need for cheap portable biosensors in medicine,pharmaceuticals, food processing and other applications. The COVID-19pandemic has accelerated the search for effective and cheap solutionsfor mass testing for infectious diseases. What is needed is accuratetesting that can be performed quickly and efficiently with minimal riskof false readings by ordinary healthcare workers. The tests could bedelivered on the doorstep or in a large batch processing settingtargeting a throughput of around 10,000 samples per day.

A major limitation of current test equipment is that many processingsteps are needed to convert the material captured by an initial nose orthroat swab sample into a material that can be analyzed using a standardinstrument. For example, steps are usually required for converting RNAmolecules to complementary DNA, a similar molecular which can be handledmore easily. The process can typically involve sample purification,making reaction mixtures and adding RNA templates. Heating steps toamplify the viral RNA are also required.

A spectrophotometric sensor is required for reading the fluorescencegenerated by the amplified RNA (amplicons). One lab-based RNA testingtechnique, Reverse-Transcription Polymerase Chain Reaction (RT-PCR),which is widely used in fields such as genetics, can detect whetherviral RNA is present by capturing and amplifying regions of the virus'genetic material. The viral RNA is converted to DNA, copied many timesuntil enough material is available for detection. The material issubjected to repeated temperature cycles then detected with the aid offluorescent markers. Processing of the sample typically takes from 3-4hours.

Another technique for viral RNA testing, Recombinase Polymerase

Amplification (RPA), is as specific as PCR amplification, but is muchfaster and does not require thermal processing. Once initiated, theamplification reaction progresses rapidly, so that starting with just afew target copies of DNA, the highly specific DNA amplification reachesdetectable levels within minutes.

Loop-Mediated Isothermal Amplification (LAMP) eliminates temperaturecycle and is fast, stable, sensitive, more specific for DNAidentification and can be integrated with microfluidics. LAMP-basedapproaches produce approx. 50-fold more amplicons than PCR-basedmethods. Importantly, LAMP can amplify NA in complex substrates even inthe presence of contaminants However, the LAMP technique is less maturethan PCR.

In general, current gold standard equipment is very expensive, costingmany thousands of dollars, and is too bulky to be brought to thepatient. These disadvantages make current equipment unsuitable fordeployment in GP surgeries, pharmacies or care homes or for thelarge-scale testing demanded by the current COVID-19 outbreak.

SUMMARY OF THE INVENTION

An object of the invention is to solve at least the above problemsand/or disadvantages and to provide at least the advantages describedhereinafter.

Therefore, an object of the present invention is to provide a generalpurpose sensor architecture integrating a surface enhanced Ramanspectroscopy (SERS) substrate, a diffractive laser beam deliverysubstrate and a diffractive infrared detection substrate.

Another object of the present invention is to provide processes forlarge volume fabrication of SERS substrates.

Another object of the present invention is to provide diffractingstructures for integrating SERS laser irradiation and Raman scattercollection, including orthogonal grating structures.

Another object of the present invention is to provide diffractingstructures for long wavelength electromagnetic radiation collection anddetector coupling.

Another object of the present invention is to provide diffractingstructures incorporating antennas and other components forelectromagnetic communications.

Another object of the present invention is to provide a gratingstructures and fabrication processes for pixel arrays and associatedelectronic signal and control circuity for use in high density liquidcrystals displays.

Another object of the present invention is to provide laser beamdirecting optical structures using switching gratings for SERSactivation.

Another object of the present invention is to provide diffractingstructures incorporating features for optimizing plasmon fieldcharacteristics in SERS devices.

Another object of the present invention is to provide SERS substratesincorporating liquid crystal layers.

Another object of the present invention is to provide SERS substratesincorporating nanostructures.

Another object of the present invention is to provide SERS substratesincorporating reporter molecules.

Another object of the present invention is to provide SERS substratesincorporating phase-separated diffractive structures.

Another object of the present invention is to provide SERS substratesincorporating metallized diffractive structures.

Another object of the present invention is to provide SERS substratesincorporating hybrid diffractive structures.

Another object of the present invention is to provide reconfigurablediffractive antennas for wireless communications, imaging and sensingapplications.

Another object of the present invention is to provide a sensorarchitecture for detecting COVID-19 from saliva.

Another object of the present invention is to provide grating structuresand grating structure fabrication processes for millimeter waves.

To achieve at least the above objects, in whole or in part, there isprovided a biosensor, comprising a pump laser source that emits pumplight having at least one wavelength, a surface enhanced Ramanspectroscopy (SERS) substrate. an analyte layer comprising at least onetype of molecule disposed on said SERS substrate, each of said at leastone type of molecule exhibiting a unique Raman spectrum underirradiation from said light at said at least one wavelength, a pumplaser substate comprising at least one pump laser channel forpropagating pump laser beams, a pump beam switch for directing a portionof the pump light from said pump laser source into each of said pumplaser channels sequentially, a coupling layer overlaying each of said atleast one pump laser channel for directing the pump light portionpropagating in each channel towards said SERS substrate, a Raman signaldetection substrate comprising at least one Raman signal detectionchannel supporting a surface relief structure for capturing a Ramansignal and guiding said Raman signal, a detector with a detectionbandwidth covering a Raman spectra of said molecules, and a Raman signalbeam combiner for coupling each of said at least one Raman signaldetection channel to said detector.

Additional advantages, objects, and features of the invention will beset forth in part in the description which follows and in part willbecome apparent to those having ordinary skill in the art uponexamination of the following or may be learned from practice of theinvention. The objects and advantages of the invention may be realizedand attained as particularly pointed out in the appended claims.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention will be described in detail with reference to thefollowing drawings in which like reference numerals refer to likeelements wherein:

FIG. 1 is a perspective view of a waveguide biosensor, in accordancewith an embodiment of the present invention;

FIG. 2 is a side elevation view of a waveguide biosensor, in accordancewith an embodiment of the present invention, in accordance with anembodiment of the present invention;

FIG. 3 is a cross-sectional view of the waveguide biosensor of FIG. 3 ;

FIG. 4 is a first plan view of components of the waveguide biosensor ofFIG. 2 ;

FIG. 5 is a second plan view of components of the waveguide biosensor ofFIG. 2 ;

FIG. 6A is a perspective view of a pump laser channel array and aswitchable grating waveguide pump beam switching module, in accordancewith an embodiment of the invention;

FIG. 6B is a plan view of a switchable grating waveguide pump beamswitching module, in accordance with an embodiment of the invention;

FIG. 7 is a side elevation view of a first example of a waveguidebiosensor in which the Raman signal channel array and the pump laserchannel array are integrated into a common waveguide structure, inaccordance with an embodiment of the invention;

FIG. 8 is a side elevation view of a first example of a waveguidebiosensor in which the Raman signal channel array and the pump laserchannel array are integrated into a common waveguide structure, inaccordance with an embodiment of the invention;

FIG. 9 is a side elevation view of a third example of a waveguidebiosensor in which the Raman signal channel array and the pump laserchannel array are integrated into a common waveguide structure areintegrated into a common waveguide structure, in accordance with anembodiment of the invention;

FIG. 10 is a side elevation view of a surface plasmon resonancesubstrate employing a noble metal layer, in accordance with anembodiment of the invention;

FIG. 11 is a side elevation view of a surface plasmon resonancesubstrate employing noble metal nanoparticles deposited onto asubstrate, in accordance with an embodiment of the invention;

FIG. 12 is a side elevation view of a surface plasmon resonancesubstrate comprising a nanostructured surface to which a noble metalcoating has been applied, in accordance with an embodiment of theinvention;

FIG. 13 is a side elevation view of a surface plasmon resonancesubstrate comprising a nanostructured surface formed using a phaseseparation process to which a noble metal coating has been applied, inaccordance with an embodiment of the invention;

FIG. 14 is a side elevation view of a surface plasmon resonancesubstrate comprising nanoparticles formed from noble metal nano shellsto which a Raman reporter molecule has been applied, in accordance withan embodiment of the invention;

FIG. 15 is a plan view of a portion of a waveguide biosensor based onthe waveguide biosensor of FIG. 2 , in which an array of beam intensitymodifying filters matched to the scattering cross sections of dominantRaman lines of a molecule to be detected are disposed between the pumplaser beam switch and the pump beam channel array, in accordance with anembodiment of the invention;

FIG. 16A is a side elevation view of a Raman signal channel employing atilted reflector surface, in accordance with an embodiment of theinvention;

FIG. 16B is a side elevation view of a Raman signal channel employing areflector surface incorporating a multiplicity of facets, in accordancewith an embodiment of the invention;

FIG. 17 is a cross-sectional view of a waveguide biosensor in which thepump laser channel array incorporates surfaces or coatings forinhibiting cross talk between adjacent channels, in accordance with anembodiment of the invention;

FIG. 18 is a cross-sectional view of a waveguide biosensor in which theRaman signal channel array incorporates surfaces or coatings forinhibiting cross talk between adjacent channels, in accordance with anembodiment of the invention;

FIG. 19A is a cross-sectional view of a biosensor that integrates asurface plasmon resonance surface with a diffractive detector lens, inaccordance with an embodiment of the invention;

FIG. 19B is a plan view of a portion of a biosensor, based on thebiosensor of FIG. 19A, that integrates a surface plasmon resonancesurface with a diffractive detector lens showing the disposition of thesurface plasmon resonance substrate/lens, the analyte layer and areflector, in accordance with an embodiment of the invention;

FIG. 19C is a cross-sectional view of a first example of an integrationof a surface plasmon resonance surface with a diffractive detector lensbased on a polymer grating structure backfilled with noble metalnanoparticles, in accordance with an embodiment of the invention;

FIG. 19D is a cross-sectional view of a first example of the integrationof a surface plasmon resonance surface with a diffractive detector lensbased on a polymer grating structure multiplexing different gratingstructures for Raman excitation and Raman signal detection, inaccordance with an embodiment of the invention;

FIG. 20 is a flowchart of a process for detecting a molecule using awaveguide sensor, in accordance with an embodiment of the invention;

FIG. 21 is a cross-sectional view of a SERS substrate including a liquidcrystal and polymer layer and a dielectric layer, in accordance with anembodiment of the invention;

FIG. 22 is a cross-sectional view of a SERS substrate including a liquidcrystal and polymer layer, in accordance with an embodiment of theinvention;

FIG. 23 is a cross-sectional view of a SERS substrate including a liquidcrystal and polymer layer formed as a nanostructure, in accordance withan embodiment of the invention;

FIG. 24 is a perspective view of a biosensor including a waveguide forpropagating a Raman signal and a waveguide for propagating pump laserbeams, in accordance with an embodiment of the invention;

FIG. 25 is a perspective view of a biosensor including a waveguidesupporting orthogonal grating structures and configured for propagatingRaman signals pump and pump laser beams in orthogonal directions, inaccordance with an embodiment of the invention;

FIG. 26 is a cross-sectional view of a portion of a waveguide supportingorthogonal grating structures on one face and a SERS element on anopposing face and configured for propagating Raman signals pump and pumplaser beams in orthogonal directions, in accordance with an embodimentof the invention;

FIG. 27 is a cross-sectional view of a portion of a waveguide supportingorthogonal grating structures and SERS elements disposed on one face ofthe waveguide and configured for propagating Raman signals pump and pumplaser beams in orthogonal directions, in accordance with an embodimentof the invention;

FIG. 28A is a cross-sectional view of a portion of a waveguidesupporting a metallic or dielectric diffracting structure applied usinga coating process showing an out-coupled ray, in accordance with anembodiment of the invention;

FIG. 28B is a cross-sectional view of a portion of a waveguidesupporting a metallic or dielectric diffracting structure applied usinga coating process showing an in-coupled ray, in accordance with anembodiment of the invention;

FIG. 28C is a cross-sectional view of a portion of a waveguidesupporting a metallic or dielectric diffracting structure applied usinga coating process configured to direct light to an analyte coated SERSsubstrate showing an out-coupled ray, in accordance with an embodimentof the invention;

FIG. 28D is a cross-sectional view of a portion of a waveguidesupporting a metallic or dielectric diffracting structure applied usinga coating process configured to direct light to an analyte coated SERSsubstrate showing an in-coupled ray, in accordance with an embodiment ofthe invention;

FIG. 28E is a cross-sectional view of a first operational mode portionof a waveguide supporting a metallic or dielectric diffracting structureapplied using a coating process and a grating formed using phaseseparation, in accordance with an embodiment of the invention;

FIG. 28F is a cross-sectional view of a second operational mode of aportion of a waveguide supporting a metallic or dielectric diffractingstructure applied using a coating process and a grating formed usingphase separation, in accordance with an embodiment of the invention;

FIG. 28G is a cross-sectional view of a first operational mode of aportion of a waveguide supporting a metallic or dielectric diffractingstructure applied using a coating process and a grating formed usingphase separation configured to direct light to an analyte coated SERSsubstrate, in accordance with an embodiment of the invention;

FIG. 28H is a cross-sectional view of a second operational mode of aportion of a waveguide supporting a metallic or dielectric diffractingstructure applied using a coating process and a grating formed usingphase separation configured to direct light to an analyte coated SERSsubstrate, in accordance with an embodiment of the invention;

FIG. 28I is a cross-sectional view of a portion of a waveguidesupporting a grating formed using phase separation with a patternedelectrode applied to the upper surface of the waveguide and anon-patterned electrode applied to the bottom surface of the waveguide,in accordance with an embodiment of the invention;

FIG. 28J is a cross-sectional view of a portion of a waveguidesupporting a grating formed using phase separation with a gratingconfigured to diffract first wavelength light applied to the bottomsurface of the waveguide and a grating patterned to diffract secondwavelength light applied to the top surface of the waveguide, inaccordance with an embodiment of the invention;

FIG. 29A illustrates a first step in the formation of phase-separatedgrating comprising depositing a liquid crystal and monomer layer onto asubstrate, in accordance with an embodiment of the invention;

FIG. 29B illustrates a second step in the formation of phase-separatedgrating comprising phase separating the liquid crystal and monomer layerinto liquid-crystal-rich and polymer-rich grating regions, in accordancewith an embodiment of the invention;

FIG. 29C illustrates a third step in the formation of phase-separatedgrating comprising removing the liquid crystal from the grating, inaccordance with an embodiment of the invention;

FIG. 29D illustrates a fourth step in the formation of phase-separatedgrating comprising metal-coating the polymer grating formed by removingthe liquid crystal, in accordance with an embodiment of the invention;

FIG. 29E illustrates a fifth step in the formation of phase-separatedgrating comprising the removal of an upper layer of the coated gratingmaterial, in accordance with an embodiment of the invention;

FIG. 29F illustrates a sixth step in the formation of phase-separatedgrating show a finished planarize composite grating structure, inaccordance with an embodiment of the invention;

FIG. 30 is a flowchart of a process for fabricating a composite polymerand metal grating structure, in accordance with an embodiment of theinvention;

FIG. 31 is a perspective view of a biosensor including a waveguide forpropagating a Raman signal and propagating pump laser beams and furthercomprising a switchable diffractive structure, in accordance with anembodiment of the invention;

FIG. 32 is a plan view of elements of an active matrix thin filmtransistor type pixel array in which the pixel electrodes and signal andcontrol circuitry are formed using a phase separation process, inaccordance with an embodiment of the invention;

FIG. 33 is a plan view of elements of an antenna comprising ananostructure incorporating a liquid crystal component, in accordancewith an embodiment of the invention;

FIG. 34 illustrates a phased separated Bragg grating structurecomprising polymer fringes immersed in air with each polymer fringecontaining nanoparticles dispersed within a polymer matrix, inaccordance with an embodiment of the invention;

FIG. 35 illustrates a slice through a three-dimensional phase-separatedphotonic crystal or lattice comprises polymer regions immersed in airwith each polymer region containing nanoparticles dispersed within apolymer matrix, in accordance with an embodiment of the invention;

FIG. 36 illustrates a phase-separated multiplexed grating structureformed from crossed Bragg gratings immersed in air, where each gratingcomprises fringes containing nanoparticles dispersed within a polymermatrix, in accordance with an embodiment of the invention; and

FIG. 37 illustrates a phase-separated diffractive structure in which theevacuated regions resulting from the evacuation of liquid crystal maycontain a residual polymer network including gold or silvernanoparticles suspended in the polymer network, in accordance with anembodiment of the invention.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

The following description is provided to enable a person of ordinaryskill in the art to make and use the invention and sets forth the bestmodes contemplated by the inventor for carrying out the invention.Various modifications, however, will remain readily apparent to thoseskilled in the art, since the general principles of the presentinvention have been defined herein specifically to provide exampleembodiments.

For the purposes of explaining the present invention, well-knownfeatures of optical technology known to those skilled in the art ofoptical design and visual displays have been omitted or simplified inorder to not obscure the basic principles of the invention. In thefollowing description, the terms “light,” “ray,” “beam” and “direction”may be used interchangeably and in association with each other toindicate the direction of propagation of electromagnetic radiation alongrectilinear trajectories. The terms “light” and “illumination” may beused in relation to the visible and infrared bands of theelectromagnetic spectrum. Parts of the following description will bepresented using terminology commonly employed by those skilled in theart of optical design. It should also be noted that in the followingdescription of the invention, repeated usage of the phrases “in oneembodiment” “an embodiment” does not necessarily refer to the sameembodiment. As used herein, the term “grating” may encompass a gratingcomprised of a set of gratings in some embodiments.

It should be appreciated that various concepts and embodiments discussedherein may be implemented in any of numerous ways, as the disclosedconcepts and embodiments are not limited to any particular manner ofimplementation. Examples of specific implementations and applicationsare provided primarily for illustrative purposes. A more completeunderstanding of the present invention can be obtained by consideringthe following detailed description in conjunction with the accompanyingdrawings, wherein like index numerals indicate like parts. For purposesof clarity, details relating to technical material that is known in thetechnical fields related to the invention have not been described indetail.

The prospect of miniaturizing biological test processes using theconcept of a lab-on-a-chip has attracted interest from many quarters.Such devices can integrate many of the functions of sample acquisition,chemical processing and flow management and can support compactgrating-based devices for performing spectrophotometric analysis. Themost well-known examples of grating sensors are holograms that canchange their color when illuminated by light scattered from molecularstructures. Miniature grating spectrophotometers for carry out moresophisticated spectrophotometric assays are also widely used in manyfields.

Gratings that can be chemically functionalized by means of dopants addedto holographic film prior to exposure, within cladding layers overlayingthe grating or onto surface relief gratings have also been widelyreported. Many of the functions required for routing samples to on-chipprocessing nodes can be implemented as microfluidics systems.

Optical waveguides can provide a compact transparent device forimplementing grating sensors. The versatility of gratings can be greatlyenhanced by using switching gratings recorded in holographic polymerdispersed material systems. U.S. Pat. No. 5,937,115, entitled“Switchable Optical Components/Structures and Methods for theFabrication Thereof,” which is incorporated herein by reference itsentirety, discloses a family of electro-optical components based onholographic polymer dispersed liquid crystal (HPDLC) gratings.

There is a need for a low cost, efficient, compact apparatus based on agrating waveguide for detecting molecular structures dispersed in asample. Such an apparatus could be used to implement a low-cost, compactlab-on-a-chip biosensor that can meet the needs of large-scaleinfectious disease testing. Such a device could also have many otherapplications in which molecules present in the liquid, gaseous or solidphases need to be characterized reliably, cost-effectively and withminimal intervention by highly skilled personnel.

Outline of the Biosensor

An object of the invention is achieved in a first embodiment in whichthere is provided a biosensor for detecting at least one type ofmolecule dispersed in at least one sample. As shown in the perspectiveview of FIG. 1 and in the cross-sectional views and plan views of FIGS.2-5 , the apparatus comprises a detector (112), a laser source (101),and a laser beam switch (102) for sequentially directing the laser beaminto a substrate supporting a multiplicity of pump laser channels (103).By switching the input beam between channels, the guided pump beams(104) in each pump beam channel can provide extracted portions (105) viaa pump beam output grating (which is shown in the side andcross-sectional views of the biosensor in FIGS. 2-3 ) for illuminating aportion of a surface plasmon resonance substrate (106) overlapping thepump beam channel.

Disposed above the surface plasmon resonance substrate (106) is a layerof analyte (107) containing a multiplicity of molecules in a layerdisposed between the surface plasmon resonance substrate (106) and theRaman signal channels. The Raman signal (108) scattered from the analyteafter stimulation by the laser beam and amplification by the surfaceplasmon resonance substrate (106) can be coupled into a furthersubstrate supporting a plurality of Raman signal channels (109), eachsandwiched by reflectors (113 and 114). Each channel is operative toin-couple and convey the Raman scatter signal (110) from the analyteand, as shown in the following figures, further comprises an upperreflector (113) and a lower reflector (114) sandwiching each Ramansignal channel (109), each Raman signal channel (109) overlapping a pumplaser channel (103). Each Raman signal channel (109) supports opticalstructures for selectively coupling in predefined portions of the Ramanspectrum emitted by the analyte (107). The required spectral selectivitycan be provided by gratings formed in one or both of the upper and lowerreflectors (113 and 114), the gratings having prescriptions optimizedfor high efficiency diffraction in said portions of the Raman spectrum.In some embodiments, optical filters can be used to filter portions ofthe Raman spectrum. The guided Raman signal in each channel is conveyedtowards a beam combiner (111) for coupling to the detector (112). Aswill be discussed below, the above layers can be configured in severaldifferent ways, including combining the pump laser channel substrate andthe Raman signal channel substrate into a single waveguiding structure.It should be apparent from FIG. 1 that a simple implementation of thepresent invention only requires one pump laser channel (103) and oneRaman signal channel (109). FIGS. 4-5 show plan views of several of theabove described elements of the biosensor.

The biosensor will now be described in more detail with reference toFIGS. 1-33 , which illustrate various aspects of the biosensor, inaccordance with many embodiments of the invention. Although, thefollowing description will concentrate on the detection of Ramanscatter, it should be apparent from the drawings and description thatthe invention can be implemented using a range of scattering phenomena,including resonant Raman scatter, non-resonant Raman scatter, CoherentAnti Stokes Raman Spectroscopy (CARS), Rayleigh scatter, Mie scatteringand Brillouin scattering. It should also be apparent that, with suitablechoice of source, detector and waveguide prescription, a biosensor basedon the principles discussed herein can be configured for a range ofsensing applications extending from the UV to radio wave bands.

Source

In some embodiments of the present invention, the source (101) emits atleast one pump wavelength for stimulating Raman scatter in a molecule.The present invention is not limited to any specific emitter technologyor operating wavelength. Many different types and configurations oflight sources can be used while still falling within the scope of thepresent invention. Possible sources include, but are not limited to, alaser, a laser array, a plurality of laser die emitting at differentwavelengths, a light emitting diode, a quantum dot, a broadband lightsource, and a broadband light source used in conjunction with a set ofspectral bandpass filters for extract narrower wavelength bands. In someembodiments, the source may comprise one or more laser devices. Lasersoffer the high wavelength specificity for molecular characterization.Lasers are also compatible with diffractive optical systems. In someembodiments, frequency tuneable lasers can be used. In some embodiments,laser arrays may be advantageous for providing parallel sampleprocessing. In some embodiments, the laser can provide a pulsed output.In some embodiments where high wavelength specificity is not a keyrequirement, broadband sources such as a light emitting diode, organiclight emitting diodes or tungsten halogen lamp can be used. In someembodiments, a broadband light source can be used in conjunction with aset of spectral bandpass filters for extracting narrower wavelengthbands. In some embodiments, quantum dots, which have peak wavelengthemission controlled by dot size, can be used to provide multiplewavelength sources. In some embodiments, the pump laser preferably emitsin the near infrared at 780 nm. In some embodiments, the pump laserpreferably emits in the ultraviolet at a wavelength less the 350 nm.

Detector

Many different types of detectors can be used in the present invention.In some embodiments, the detector (112) can be a single element device.In some embodiments, the detector (112) can be an array ofphotosensitive elements. In some embodiments, the detector (112) ispreferably configured to operate over Raman signal spectral bandwidth.In some embodiments, the detector (112) preferably operates over theMWIR wavelength range. In some embodiments, a signal processor ispreferably connected to the detector (112). In some embodiments, theRaman signal comprises an intensity variation across a wavelength band.

Pump Laser Channels

In some embodiments, a plurality of pump laser channels (103) can besupported by a substrate. In some embodiments, the substrate is atransparent optical material such as glass or an optical plastic. Insome embodiments, the substate supports a coupling grating forextracting light from the pump laser channels towards the surfaceplasmon resonance substrate. In some embodiments, the coupling gratingcan have a prescription for directing the wavevectors of the diffractedpump laser beams to satisfy Raman scatter momentum balance conditionswithing the analyte and surface plasmon interaction region. In someembodiments, the pump laser channels (103) can be configured to interactwith at least one layer for modify the pump laser beam polarization. Insome embodiments, it is important to avoid leakage of light from onechannel to another. In some embodiments, the pump laser channels (103)can be formed by core materials embedded within the substrate where thecore refractive index is higher than the substrate refractive index. Insome embodiments, the pump laser channels (103) can be formed as ridgestructures disposed on a substrate surface.

In some embodiments, such as the one illustrated in FIGS. 17-18 , therequired confinement of the pump laser beams can be provided usingcoatings (129) designed to scatter, reflect or absorb light that mightotherwise leak into an adjacent channel. The coatings (129) can beapplied to substate surface portions on either side of each channel. Insome embodiments, coatings or surface etched features can be applied tothe substrates surface portions overlapping each channel. The advantageof this technique is that the need for etching or diffusion of waveguidecores into the substrate can be avoided Raman signals typically exhibitforward and backward scattered intensity lobes. In some embodiments, areflector can be disposed under the pump laser channel array substrateto reflect backward-scattered lobes of the Raman scatter distributiontowards the Raman signal channel array. In some embodiments, a pumplaser channel (103) can be provided by a solid or hollow light pipehaving a continuous or polygonal cross-section. In some embodiments, apump laser channel (103) can further comprise a light trap for capturingzero order diffracted light.

Switchable Beam Directing Module

In some embodiments, a switchable beam directing module (102) is used todirect light from the source (101) into TIR paths within each of thepump laser channels sequentially. The beam directing module (102) can beconfigured in many ways. In some embodiments, a plurality of laser beamscan be sequentially generated using arrays of switchable gratings. Onesolution, illustrated in FIGS. 6A-6B, advantageously uses a waveguidecontaining switchable grating elements. FIG. 6A illustrates a portion ofthe pump laser channel array (103), in which each channel is sandwichedby isolation regions for avoiding crosstalk, and a pump laser beamswitch (102) coupled to a pump laser source (101) emitting a pump laserbeam (101A) which can be coupled into total internal reflection pathsinside a waveguide structure (102A) comprising substrates sandwiching aswitching grating layer (102B) into which are formed a linear array ofswitching grating elements (102C). In many embodiments, the apparatuscan further comprise a beam stop (102D). Seven elements are illustrated.However, in some embodiments more elements or fewer elements can beused. Each grating element when switched from its passive state into itsdiffracting state diffracts light out of the waveguide structure intoone of the pump laser channels (103).

FIG. 6B is a plan view of the waveguiding structure. Each gratingelement has a grating vector (or K-vector) for deflecting a laser beamundergoing total internal reflection within the waveguide out of thewaveguide at 90 degrees to the waveguide surface. Each grating elementin its diffracting state directs the laser beam into one of the pumplaser channels (103). In some embodiments, the switchable gratings(102C) can be configured as transmission gratings. In some embodiments,the switchable gratings (102C) can operate as reflection gratings. Sincethe lasers for stimulating Raman scatter tend to emit in the visible tonear infrared bands, the switchable gratings (102C) can be providedusing polymer and liquid crystal material systems.

FIG. 15 is a plan view of a portion of a waveguide biosensor in which anarray of beam intensity modifying filters (127) matched to thescattering cross sections of dominant Raman lines of a molecule to bedetected are disposed between the pump laser beam switch (102) and thepump beam channel array, in accordance with an embodiment of theinvention.

Surface Plasmon Resonance Substrate

The surface plasmon resonance substrate is used for amplifying at leastone of said laser light and the Raman scatter. Examples of surfaceplasmon resonance substrates that can be used in the present inventionare illustrated in FIGS. 10-13 . In some embodiments, such as theembodiment shown in FIG. 10 , the surface plasmon resonance substrate issuitably a noble metal coating (116), such as gold, silver, oraluminium, applied to an optical substrate. In some embodiments, thesurface plasmon resonance substrate can be provided by a phase-separatedholographic polymer nanocomposites using noble metallic nanoparticles(gold, silver etc.). In some embodiments, the surface plasmon resonancesubstrate can be provided by a polymer surface nanostructured gratingformed on an optical substrate and backfilled with metal nanoparticle.In some embodiments, the surface plasmon resonance substrate can beprovided by noble metal nanoparticles supported by an optical substate.In some embodiments, the surface plasmon resonance substrate can beprovided by a nanostructured metallic surface. In some embodiments, thenanostructure surfaces can be fabricated using Rolling Mask Lithography(RML).

FIG. 11 shows a surface plasmon resonance surface comprising gold orsilver nanoparticles (117) dispersed within a supporting medium (118).FIG. 12 shows a surface plasmon resonance surface formed from asubstrate with a nanostructured surface (119) coated with a gold orsilver film (120). FIG. 13 shows a surface plasmon resonance surfaceformed using a phase separation process resulting in a nanostructuredsurface (121) coated with a gold or silver film (120). In someembodiments, the nanostructured surface (121) can be patterned. In someembodiments, the patterning can be designed to facilitate surfaceplasmon intensity distributions matched to a molecule to be analyzed. Insome embodiments, the surface plasmon resonance substrate can beprovided by a shell of a nanoparticle. In some embodiments, the surfaceplasmon resonance substrate can be provided by multiplicities ofplasmonic-magnetic silica nanotubes supported by an optical substrate.In some embodiments, the surface plasmon resonance substrate can beprovided by a substrates supporting layers comprising a noble metalcoating for forming surface plasmons and a Raman reporter moleculecoating.

In some embodiments, the surface plasmon resonance substrate can beprovided by a substrates supporting a multiplicity of nanoparticles ornanostructures formed on said substrate with shell structures formedfrom layer comprising a noble metal coating and a Raman molecule coatingapplied to the exterior of the shell. FIG. 14 illustrates one suchembodiment in which tagged nanoparticles (123) are dispersed within asupporting medium (122). Each tagged nanoparticle (123) comprises a coreprovided by a void or inert material (124) surrounded by a gold orsilver nano shell (125) which is in turn surrounded by a Raman reportermolecule coating (126). In some embodiments, a multiplicity of opticallypowered elements can be disposed in a layer overlapping the surfaceplasmon resonance substrate for concentrating the Raman signal fromsurface plasmon hot spots formed by the surface plasmon resonancesubstrate. In some embodiments, the surface plasmon resonance substrateincorporates nanostructures for diffracting Raman scattered light. Insome embodiments, the surface plasmon resonance substrate incorporatesnanostructures forming a diffractive lens or mirror.

Raman Signal Channels

A plurality of Raman signal channels can be formed in or on a substrate,with each channel operative to in-couple and convey Raman scatter andfurther comprising an upper reflector and a lower reflector sandwichingsaid Raman signal channels, each Raman signal channel overlapping a pumplaser channel. Each Raman signal channel supports optical structureswhich, in some embodiments, are formed in one or both of the upper andlower reflectors, for selectively coupling in predefined portions of theRaman spectrum emitted by the analyte. The required spectral selectivitycan be provided by gratings formed in one or both of the upper and lowerreflectors, the gratings having prescriptions optimized for highefficiency diffraction in said portions of the Raman spectrum. In someembodiments, optical filters can be used to filter portions of the Ramanspectrum.

In some embodiments, the reflectors can be configured as at least oneselected from: a diffractive surface; a Fresnel structure; a chirpeddiffracting or reflecting structure; a surface having a spatialvariation of at least one of diffraction efficiency; reflectioncoefficient and transmission coefficient; a structure formed on aninclined surface; a glancing incidence surface; a birdbath reflector; astructure formed on a curved surface; and a surface coated with at leastone film providing selective transmission, absorption or reflection atone or more wavelengths within the spectral bandwidth of the Ramansignal.

FIG. 16A illustrates a Raman signal channel employing a tilted reflectorsurface, in accordance with an embodiment of invention. The channelcomprises a tilted upper reflector (113A) and a lower reflector (114)sandwiching the optical medium (128) for propagating the Raman signal.In some embodiments, the Raman signal channels can contain amultiplicity of tilted partially reflecting mirrors for in couplingRaman signals from the analyte. FIG. 16B illustrates a Raman signalchannel employing a facetted reflector surface (113B), in accordancewith an embodiment of invention. A detail of the reflector surfaceillustrating the reflector substrate (113C) and the facet structure(113D) is inset in FIG. 16B. In some embodiments, a Raman signal channelcan be provided by a solid or hollow light pipe having a continuous orpolygonal cross-section. In some embodiments, a Raman signal channel canfurther comprise a light trap for capturing zero order diffracted light.

In many embodiments the signal coupled into the Raman signal channel canbe confined vertically by upper and lower reflectors. The upper andlower reflectors can have grating structures coated with a metallic filmto provide high efficiency reflection for light in predefined wavelengthband. In some embodiments, each channel can have a unique range ofK-vector directions tuned to diffract a predefined wavelength band. Insome embodiments, the wavelength bands can be specified to windowprominent lines in the Raman spectrum of a molecule.

As in the case of the pump laser channel array, it is important to avoidleakage of light from one channel to another. In some embodiments, theRaman signal channels can be formed by core materials embeded within thesubstrate, where the core refractive index is higher than the substraterefractive index. In some embodiments, the Raman signal laser channelscan be formed as ridge structures disposed on a substrate surface.

In some embodiments, such as the one illustrated in FIG. 18 , therequired confinement of the Raman signal beams can be provided usingcoatings (129) designed to scatter, reflect or absorb light that mightotherwise leak into an adjacent channel The coatings (129) can beapplied to substate surface portions on either side of each channel. Insome embodiments, coatings or surface etched features can be applied tothe substrate's surface portions overlapping each channel. The advantageof this technique is that the need for etching or diffusion of waveguidecores into the substrate can be avoided.

Raman Signal Combiner

In some embodiments, a Raman signal combiner is provided for opticallycoupling the Raman signal channels to an optical path leading to thedetector. In some embodiments, the Raman signal combiner can employswitchable gratings to eliminated cross talk In some embodiments, theRaman signal combiner can incorporate bandpass or high-pass/lo-passfilters. In some embodiments, the Raman signal combiner can includeoptical components for improving signal to noise ratio, such as, forexample, polarization control elements.

Configurations of the Pump Laser Channels and Raman Signal Channels

In some embodiments, the plurality of pump laser channels and theplurality of Raman signal channels can sandwich the surface plasmonresonance substrate. In some embodiments, the plurality of pump laserchannels and the plurality of Raman signal channels can be disposedabove or below the surface plasmon resonance substrate. In someembodiments, the plurality of pump laser channels and the plurality ofRaman signal channels can be combined in a single waveguide structure inwhich the Raman signal and the pump laser beams propagate in the samedirection within the single waveguide structure, the source and theswitchable beam directing module are disposed at one end of the singlewaveguide structure, and the Raman signal combiner and the detector aredisposed at the opposite end of the single waveguide structure.

As illustrated in FIGS. 7-9 , in some embodiments, of which the keycomponents have already been discussed above, the plurality of pumplaser channels and the plurality of Raman signal channels can becombined in a single waveguide structure in which the Raman signal andthe pump laser beams counter propagate in opposing directions, thesingle waveguide structure and the source and the switchable beamdirecting module are dispose at one end of the single waveguidestructure, and the Raman signal combiner and the detector are disposedat the same end of the single waveguide structure.

Types of Gratings Used

As is apparent from the above discussion, gratings can play several keyroles in the biosensor of the present invention. Gratings can be usedfor coupling pump beams in the surface plasmon resonance substrate, andfor coupling and confining light within the Raman signal channel array.Switching gratings can be used to couple laser beams from the sourceinto each of the pump laser channels sequentially.

The switchable grating used in the present invention is preferably aBragg grating (also referred to as a volume grating). Bragg gratingshave high efficiency with little light being diffracted into higherorders. The relative amount of light in the diffracted and zero ordercan be varied by controlling the refractive index modulation of thegrating, a property which is used to make lossy waveguide gratings forextracting light over a large pupil.

One important class of gratings is known as Switchable Bragg Gratings(SBG). SBGs are fabricated by first placing a thin film of a mixture ofphotopolymerizable monomers and liquid crystal material between parallelglass plates. One or both glass plates support electrodes, typicallytransparent indium tin oxide films, for applying an electric fieldacross the film A volume phase grating is then recorded by illuminatingthe liquid material with two mutually coherent laser beams, whichinterfere to form a slanted fringe grating structure. During therecording process, the monomers polymerize, and the mixture undergoes aphase separation, creating regions densely populated by liquid crystalmicro-droplets, interspersed with regions of clear polymer. Thealternating liquid crystal-rich and liquid crystal-depleted regions formthe fringe planes of the grating. The resulting volume phase grating canexhibit very high diffraction efficiency, which may be controlled by themagnitude of the electric field applied across the film. When anelectric field is applied to the grating via transparent electrodes, thenatural orientation of the LC droplets is changed, causing therefractive index modulation of the fringes to reduce and the hologramdiffraction efficiency to drop to very low levels. Typically, SBGElements are switched clear in 30 microseconds, with a longer relaxationtime to switch ON. Note that the diffraction efficiency of the devicecan be adjusted, by means of the applied voltage, over a continuousrange. The device exhibits near 100% efficiency with no voltage appliedand essentially zero efficiency with a sufficiently high voltageapplied. In certain types of HPDLC devices, magnetic fields may be usedto control the LC orientation. In certain types of HPDLC, phaseseparation of the LC material from the polymer may be accomplished tosuch a degree that no discernible droplet structure results. An SBG mayalso be used as a passive grating. In this mode its chief benefit is auniquely high refractive index modulation.

SBGs may be used to provide transmission or reflection gratings for freespace applications. SBGs may be implemented as waveguide devices, inwhich the HPDLC forms either the waveguide core or an evanescentlycoupled layer in proximity to the waveguide. The parallel glass platesused to form the HPDLC cell provide a total internal reflection (TIR)light guiding structure. Light is coupled out of the SBG when theswitchable grating diffracts the light at an angle beyond the TIRcondition. Typically, the HPDLC used in SBGs comprise liquid crystal(LC), monomers, photoinitiator dyes, and coinitiators. The mixturefrequently includes a surfactant. The patent and scientific literaturecontains many examples of material systems and processes that may beused to fabricate SBGs. Two fundamental patents are: U.S. Pat. No.5,942,157 by Sutherland, and U.S. Pat. No. 5,751,452 by Tanaka et al.which are incorporated herein by reference in their entireties for allpurposes. Both patents describe monomer and liquid crystal materialcombinations suitable for fabricating SBG devices. One of the knownattributes of transmission SBGs is that the LC molecules tend to alignnormal to the grating fringe planes. The effect of the LC moleculealignment is that transmission SBGs efficiently diffract P-polarizedlight (i.e., light with the polarization vector in the plane ofincidence) but have nearly zero diffraction efficiency for S-polarizedlight (i.e., light with the polarization vector normal to the plane ofincidence).

Many different grating types and configuration can be used to providethe detector grating. In some embodiments the grating can be a surfacerelief grating. Typically, such gratings are formed in etchingprocesses. Surface relief gratings can also be formed by first recordinga HPDLC grating and then flushing the liquid crystal component. In someembodiments, the detector grating can be a switchable grating (a gratingelectrically switchable between two diffracting states). In someembodiments, a switchable grating can be formed in a liquid crystal andpolymer material system by phase separation of the monomer and liquidcrystal. In some embodiments, one of the diffracting states correspondsto the grating diffracting with high efficiency and the othercorresponds to the grating not diffracting and essentially acting like atransparent optical layer. In some embodiments, the grating can beswitched between diffracting states using transverse fields provided byelectrodes sandwich the grating. Other electrode schemes can be used.

In some embodiments, the grating can be switched between diffractingstates using in-plane electric fields provided by interdigitatedelectrodes. In some embodiments, a switchable detector grating can beconfigured as an array of switchable grating elements. In someembodiments, the detector grating does not need to switch. Non-switchinggratings recorded in liquid crystal and polymer material would benefitfrom exceptionally high refractive index modulations. It should be notedthat although the figures illustrate transmission gratings, equivalentembodiments can also use reflection gratings. In some embodiments, theswitching grating can be formed from a mixture including nano particles.Nanoparticles can be used to improve the electro-optical performance ofthe grating, in particular the switching time and switching voltage. Insome embodiments, the switchable grating can be formed from a mixtureincluding nanoparticles that can participate in chemical reaction withthe sample.

In various embodiments, a switchable grating provided by at least oneselected from the group of: a grating recorded in liquid crystal andpolymer material system; a grating formed by the phase separation of LCand monomer; a grating formed from a mixture comprising nano particle; agrating formed from a mixture comprising magnetic nano particles; agrating formed from a mixture comprising nanoparticles that canparticipate in chemical reaction with the sample, backfilled a passivegrating; a grating recorded in a holographic photopolymer; a surfacerelief grating; a grating formed by phase separation of LC and monomerand the flushing out of the final LC component of the cured grating; agrating recorded in a layer of recording material deposited using aninkjet coating process; a grating recorded in a layer of recordingmaterial using scanned laser illumination process; a grating recorded bycontact copy from a master; and a grating formed using a maskingprocess.

In some embodiments, a grating can be recorded in a layer of recordingmaterial deposited using an inkjet coating process. In some embodiments,the detector grating can be recorded in a layer of recording materialusing a scanned laser illumination process. In some embodiments, thedetector grating can be contact copied from a master grating. In someembodiments, masking may be used as part of the grating fabricationprocess.

In some embodiments, nanoparticles can be introduced into the grating atsome stage during grating formation or after the grating has been cured.In some embodiments, nanoparticles can fill grating voids ormicrocavities naturally occurring in a polymer grating as a result ofthe covalent single carbon bonds characteristic of the cured polymerbeing approximately 50% shorter than the Van der Waals bonds occurringin the liquid monomer state. As will be discussed below, in someembodiments, the nanoparticles can be magnetic nanoparticles. In someembodiments, the grating can incorporate molecular dopants that canreact with a molecule in the analyte to enable its identification fromthe optical signature. In some embodiments, the molecular dopants formpart of the mixture of reactants used to form the grating in a phaseseparation process. In some embodiments, the molecular dopants can bebackfilled into a surface relief grating. In some embodiments, themolecular dopants can be backfilled into a surface relief grating formedby a phase separation process. In some embodiments, the moleculardopants can be deposited within a layer overlaying the grating and lyingwithin the optical interaction zone.

In some embodiments, a grating can be configured as at least oneselected from: a chirped grating; a surface relief grating; a blazedgrating, a grating with spatially varying properties; a switchablegrating; an array of switchable grating elements; a grating configuredto interact with a microfluidic device; a grating configured to providewavelength-specific light extraction; a grating that can be switchedbetween a first diffracting state and a second diffracting state usingin plane fields provided by interdigitated electrodes; and a gratingthat can be switched between a first diffracting state and a seconddiffracting state using transverse fields provided by electrodessandwich said grating. In some embodiments, a grating can provide atleast one function selected from the group of: polarization selectivity;polarization rotation; dynamic gain equalization; wavelengthmultiplexing; variable optical attenuation; spectral filtering; beamdeflection; beam geometry shaping; beam focusing; coupling of light fromsaid source into said waveguide; and coupling of light out of saidwaveguide on to said detector.

An Embodiment Employing an Integrated Surface Plasmon ResonanceSubstrate and Diffractive Detector Lens

In some embodiments, several of the subsystem components discussed abovecan be used to provide a biosensor that uses a nanostructure substratethat provides a surface plasmon resonance substrate, while at the sametime encoding a diffractive optical element for focusing the Ramansignal onto a detector operating over the Raman signal spectralbandwidth. As in the embodiments discussed above, the optical structurescan be partitioned into regions having diffractive properties tuned tospecific wavelength bands.

In some embodiments, the apparatus comprises a laser emitting at leastone pump wavelength for stimulating Raman scatter in molecularstructure, a surface plasmon resonance substrate supportingnanostructures for forming surface plasmons integrated with ananostructure that provides a diffractive optical element for focusingthe Raman signal onto the detector, and a reflector (130). FIG. 19Aillustrates a biosensor that integrates a surface plasmon resonancesurface with a diffractive optical element (DOE) detector lens in anoptical layer (131), in accordance with an embodiment of invention. FIG.19B illustrates a portion of a biosensor, based on the principles ofFIG. 19A (integrating a surface plasmon resonance surface with adiffractive detector lens) showing the disposition of the surfaceplasmon resonance substrate/lens (131), the analyte layer and areflector (130), in accordance with an embodiment of invention.

In some embodiments, the DOE can incorporate a switchable grating arrayhaving a spatial frequency matched to the laser wavelength for couplingthe laser light at points distributed over a surface. The surfaceplasmon resonance nanostructures amplify at least one of the pump laserlight and the Raman scattered signal. In some embodiments, the surfaceplasmon resonance substrate/lens can be divided into DOE stripes, eachoperative to diffract a predefined wavelength band of the Raman signal.In some embodiments, the reflector can be divided into stripes withoptical prescriptions for diffracting a predefined band of the Ramansignal. In some embodiments, the stripe functions as a cylindrical lensor mirror. In some embodiments the DOE stripes can overlap the reflectorstripes. In some embodiments, the elements of the switchable gratingarray can be stripes that overlap the reflector stripes. In someembodiments, other grating element shapes can be used as an alternativeto stripes. For example, in some embodiments grating elements configuredas two-dimensional arrays can be used. In some embodiments, thereflector stripes can be formed at least in part from switchinggratings. In some embodiments, the surface plasmon resonance substratescan be formed using the techniques discussed above.

FIG. 19C illustrates a first example (133) of the integration of asurface plasmon resonance surface with a diffractive detector lens basedon a polymer grating structure (134) backfilled with noble metalnanoparticles (135), in accordance with an embodiment of invention. FIG.19D illustrates a first second example (136) of the integration of asurface plasmon resonance surface with a diffractive detector lens basedon a polymer grating structure (137). In some embodiments, the gratingstructure can include high resolution portions (138). In someembodiments, the grating structure is overcoated with a gold film (139)multiplexing different grating structures for Raman excitation and Ramansignal detection, in accordance with an embodiment of invention. In someembodiments, a cavity for insertion of a substrate supported withanalytes can be provided. In some embodiments, the cavity can beconfigured to be bounded by the plasmonic resonance surface and thereflection surface.

Detection Process

FIG. 20 is a flowchart of steps in a method of detecting and identifyinga molecule dispersed within a sample, in accordance with an embodimentof the invention.

As shown, a method 140 of detecting a molecule is provided. Referring tothe flowchart, method 140 includes the steps of:

-   -   a) providing (141) a source emitting at least one pump        wavelength for stimulating Raman scatter in said molecule; a        detector operating over the Raman signal spectral bandwidth; an        array of pump laser channels supported by a substrate also        supporting a coupling grating; a switchable beam directing        module for directing light from the source into TIR paths into        each of said pump laser channels sequentially; a surface plasmon        resonance substrate for amplifying at least one of said laser        light and the Raman scatter; a substate supporting a plurality        of Raman signal channels each channel operative to in couple and        convey Raman scatter and further comprising an upper reflector        and a lower reflector sandwiching said Raman signal channels,        each Raman signal channel overlapping a pump laser channel; and        a Raman signal combiner for coupling said Raman signal channels        into an optical path leading to said detector;    -   b) presenting (142) a layer of analyte containing a multiplicity        of said molecules between said surface plasmon resonance        substrate and said Raman signal channels;    -   c) directing (143) light from said source into a first pump        laser channel using said switchable beam directing module;    -   d) coupling (144) light from said first pump laser channel        towards said surface plasmon resonance substrate using said        coupling grating;    -   e) coupling (145) the Raman scatter signal from said analyte        into a first Raman signal channel overlaying said first pump        laser channel;    -   f) conveying (146) said Raman signal along said first Raman        signal channel by means of said first and second reflectors;    -   g) directing (147) said Raman signal onto said detector by means        of said Raman signal combiner;    -   h) recording (148) the Raman signal at said detector; and    -   i) measuring (149) an optical signature characterizing said        molecule

Presentation of the Analyte

In some embodiments, the analyte is in the liquid phase. However, thepresent invention can also be applied in cases were an analyte is in thegaseous or solid phase, or in a multiphase system involving two or moreof the above phases. In some embodiments, the analyte can comprisemolecules suspended in a liquid. In some embodiments, the analyte cancomprise molecules dissolved in a liquid. In some embodiments, ananalyte can comprise molecules adhering to the surfaces ofnanoparticles. In some embodiments, an analyte can comprise moleculesabsorbed within nanoparticles. In some embodiments, an analyte cancomprise molecules that can react with other marker molecules to providedetectable molecular species. In yet some embodiments, an analyte samplecan comprise molecules existing in the gaseous phase. In someembodiments, the analyte can have at least one surface in contact withan air gap. In some embodiments, the analyte can be in contact with asurface of the surface plasmon resonance substrate. In some embodiments,the analyte fills voids in a nanostructured surface formed by thesurface plasmon resonance substrate. In some embodiments, the biosensorcan further comprise a cavity for insertion of a substrate or cartridgefor supporting an analyte bounded by the surface plasmon resonancesurface and the Raman signal channel substate.

The biosensor can, in some embodiments, incorporate elements forsubjecting the analyte to a stimulus comprising at least one of:pressure, vibration, an electromagnetic field, and temperature. In someembodiments, the phenomenon of evanescent coupling may be used toimprove the efficiency of interaction between the pump laser and thesurface plasmon resonance structure and the analyte. In someembodiments, evanescent coupling takes place between the pump laserchannels and the analyte.

Analyte Delivery Systems

Although the present invention is primarily directed at the detection ofmolecules, it can also be used in conjunction with delivery of ananalyte to the detector. An “optical interaction zone” essentiallydefines a volume from which Raman scatter from the analyte can interactwith the pump laser beams and surface plasmons. The optical interactionzone can also refer to the volume of analyte that interacts with thesurface plasmons resonance substrate. In some embodiments, the opticalinteraction zone characterizes the spatial limit of photon transferbetween the analyte and one or both of the pump laser channel array orthe Raman signal channel array by means of diffractive coupling. In someembodiments, the optical interaction zone characterizes the spatiallimit of photon transfer between the analyte and one or both of the pumplaser channel array or the Raman signal channel array by means ofevanescent coupling.

In some embodiments, there is provided a sample conveyance means fortransporting the analyte from an analyte injection node to an opticalinteraction zone in proximity to the surface plasmon resonancesubstrate. In some embodiments, the sample conveyance means can includeat least one selected from the group of: electromagnets configured forapplying a magnetic field across said analyte; a microfluidic componentfor control and manipulation of said analyte; an injection node foradding nanoparticles to said analyte; an electrode structure forapplying heat to said analyte; an electrode structure for applying anelectrical stimulus to said analyte; and means for flushing the samplefrom an optical interaction zone of the apparatus.

In some embodiments, the sample conveyance means can be implemented on aseparate substrate overlaying the waveguide. However, in someembodiments, the sample conveyance means can be supported by one of thebiosensor substrates discussed above. In some embodiments, the sampleconveyance means can be based on providing an assembly of nanoparticlesto which sample molecules can adhere. In some embodiments, the samplemolecules can be fluorescently labelled biomarkers. The nanoparticlescan, in some embodiments, have magnetic properties allowing theconveyance and manipulation of the sample to be carried out usingmagnetic fields. In some embodiments, acoustic fields can be used forthe conveyance and manipulation of the sample.

In some embodiments, the sample conveyance means can be based onmagnetic particle-based target molecule extraction and transportationmethods. In some embodiments, the latter can be implemented withindisposable cartridges. Magnetic nanoparticles, such as thosemanufactured by Nanopartz Inc. (CO), are paramagnetic highlymonodisperse spherical plasmonic magnetic nanoparticles available incontrolled sizes (typically with diameters from 2 nm to 250 nm) andhighly monodisperse shapes. Magnetic nanoparticles can provide a verydense magnetic lattice resulting in a very high magnetic response.Properly engineered magnetic nanoparticles can be extremely magnetic andcan maintain homogeneous dispersion without aggregation. Magneticnanoparticles particles facilitate molecular detection by enablingplasmon resonance, wavelength-selective absorption, scattering and otheroptical properties.

In some embodiments, magnetic nanoparticles can be deposited on thedetector grating. In some embodiments, magnetic nanoparticles can bedeposited on a clad layer overlaying the grating. In some embodiments,magnetic nanoparticles can be integrated into the detector gratingrecording material. In some embodiments, magnetic nanoparticles can bedispersed in the molecular sample to be analyzed. In some embodiments,an external magnetic field can be used to direct particles alongchannels towards the optical interaction zone of the grating detector.In some embodiments, external magnetic fields can be used for theredispersion of sample molecules at stages in the conveyance of thesample from the sample injection node to the detector grating. In someembodiments, external magnetic fields can be used to separate differenttypes of molecules. In some embodiments, the sample conveyance means canfurther include electromagnets configured for a applying a magneticfield across a channel In some embodiments, the sample conveyance meanscan include one or more microfluidic components for the control,manipulation and conveyance of the sample. In some embodiments, thesample conveyance means can have one or more injection nodes for addingnanoparticles to the sample. In some embodiments, the sample conveyancemeans can incorporate an electrode structure for applying heat to thesample. In some embodiments, electrode structure can be used forapplying an electrical stimulus to the sample. In embodiments where thesample is temporarily in contact with a detector grating, means forflushing the sample from the optical interaction zone of the grating canbe provided. In some embodiments, the sample conveyance means includes acassette containing the sample which can be moved to the detectorgrating using an electro-mechanical drive a magneto-mechanical drive orby some other means. In some embodiments, a patterned transportpropulsion system can be provided. In some embodiments, the patternedtransport propulsion systems can be fabricated using 2D printingtechniques. In some embodiments, where the viscosity of the samplematerial may not be compatible with microfluidics the sample conveyancemeans may be based on applying a sample to a substrate. In someembodiments, the biosensor may include a means for freeze drying thesample. In some embodiments, the sample substates can be transportedfrom the sample injection node to the detector gratings usingelectro-mechanical or magneto-mechanical drives. In some embodiments,the sample molecules can be applied to magnetized nanoparticlescontained within a cassette. In such embodiments, the cassette can beconfigured to pass between electromagnets disposed at a location along apropulsion track. In some embodiments the formation of a suspendedassembly of sample-doped nanoparticles within a cassette can be carriedout using laser stimuli applied to the cassette. In some embodiments,the formation of a suspended assembly of molecules applied tonanoparticles within a cassette can be carried out using acoustic orvibratory stimuli applied to the cassette. In some embodiments, theformation of a directed assembly or lattice of nanoparticle to whichsample molecules can adhere can be formed by an acoustic standing waveformed within the cassette using a transducer coupled to the cassette.In some embodiments, the sample molecules can be fluorescently labelledbiomarkers. In some embodiments, the acoustic standing wave can be asurface acoustic wave formed on substrate provided inside the cassette.In some embodiments, such a nanoparticle lattice formed by the abovemeans can play the role of a detector grating. In some embodiments,acoustic waves can be used to separate different types of sample-dopedmolecules. In some embodiments, acoustic waves can be used to conveysample-doped nanoparticles along channels. In some embodiments, acousticwaves can be used to concentrate sample-doped nanoparticles to enhancethe detected signal from the sample. In some embodiments, theconcentration can be provided by acoustic focusing. In some embodiments,the concentration can be provided by vortex formation.

Other Features of the Biosensor

Many well-known techniques for improving signal-to-noise ratio andeliminating cross and stray light in optical systems can be used toimprove the performance of the biosensor. In some embodiments, theapparatus can further include at least one selected from: anelectrically variable refractive index medium; a dichroic filter, anapodizing filter and a MEMs device; an infrared absorption medium; apolarization filter; a polarization control component; a lightintegration element; and a light trap.

Applications of the Biosensor

Although the biosensor of the present invention will be discussed inrelation to the detection of infectious diseases, it should beappreciated that the systems and methods disclosed herein have manyother applications in many different fields such as, for example,biological research, industrial and agricultural process monitoring,pollution monitoring and others. The biosensor of the present inventioncan also be applied in multi-sensor systems, for example, for detectinggas, water, chemicals, heat etc. The biosensor can also be configuredfor use in wireless devices for medical diagnostics and monitoring, suchas a non-invasive consumer glucose monitor currently being developed byMediWise Ltd. which operates in the 40 GHz (radio wave) band.

Augmentation of Surface Plasmonic Nanostructures Using Liquid CrystalSystems

In some embodiments, a reconfigurable surface plasmonic resonancesubstrate can combine plasmonic nanostructures with liquid crystals,enabling surface plasmon resonance substrates that can be cheaplyreconfigured for detecting specific molecules. Surface plasmon andliquid crystals can also, in some embodiments, be used for spatiallyand/or temporally varying the characteristic of surface plasmons. Insome embodiments, patterns of plasmon hot spots can be provided in thisway. Current surface plasmon resonance substrates are expensive tomanufacture or reconfigure for different applications. Top-downnanofabrication techniques such as electron beam lithography and focusedion beam milling allow the accurate fabrication of structures atnanoscale but suffer from high capital equipment expenses, and slowfabrication cycles. Bottom-up techniques like self-assembly can easilyachieve regular patterns at a large scale at a lower cost but, in mostcases, cannot be fabricated with nanometer precision. Even small changesto a nanostructure specification will require a completely new device.There is, therefore, a requirement for reconfigurable or activeplasmonic devices. Many active mediums have been used to build activeplasmonic devices, including liquid crystals molecular machines, elasticpolymers, and chemical oxidation/reduction. Liquid crystal stands out asthe best candidate. Liquid crystals possess the smallest elasticconstants and the largest birefringence, spanning the visible toinfrared bands and beyond, among all known materials. Liquid crystalscan be chemically synthesized and processed on a very large scale, andthey are also compatible with almost all technologically importantoptoelectronic materials. Liquid crystals can be controlled byelectricity, light, acoustic waves, and other means. Efficient andversatile drive and switching techniques have been developed over theyears. By integrating liquid crystals with plasmonic nanostructures,active plasmonic materials and devices with enhanced performance havebeen demonstrated.

As discussed above, liquid crystal can be combined with polymers toenable high index modulation switching, spatio-temporally varyingrefractive index modulation and versatile means for integrating complexdiffractive optical devices, including Bragg gratings. In someembodiments, the liquid crystal polymer devices can be augmented bynanoparticles to provide the electro-optical properties. In someembodiments, the liquid crystal can be formed in a layer applied to asurface of the surface plasmon resonance substrate. In some embodiments,the liquid crystal can exist as a polymer and liquid crystal structuresforming the dielectric layer of the surface plasmon resonance substrate.

Surface plasmons are a special kind of light formed by collectiveelectron oscillations at the interface of a noble metal and dielectric.They can only propagate at the metal/dielectric interface and decayexponentially away from the interface. Conversion of propagatingelectromagnetic waves into surface plasmons requires careful matching ofthe wavevectors of the surfaces plasmons and the incident light.

In some embodiments, a liquid crystal and polymer diffractive opticalelement can be used to control the wavevectors of the incident light.FIG. 21 illustrates a surface plasmon resonance substrate (106)comprising a dielectric layer (151), a noble metal layer and a liquidcrystal and polymer layer (150). In some embodiments, the liquid crystaland polymer layer can have a dielectric constant that that enablessurface plasmon formation. FIG. 22 illustrates a surface plasmonresonance substrate comprising a noble metal layer and a polymer andliquid crystal composite layer (150). In some embodiments, a liquidcrystal and polymer layer can provide a nanostructure (FIG. 23 ) forcontrolling the spatial and temporal characteristics of surface plasmonhotspots. FIG. 23 illustrates a surface plasmon resonance substratecomprising a noble metal layer and a polymer and liquid crystalcomposite layer configured as a nanostructure (150A). The dielectricconstants of the metal and dielectric must be compatible for efficientsurface plasmon generation. Another role for a liquid crystal andpolymer layers in many embodiments can be to fine tune the relativedielectric constants at the metal and dielectric interface.

In some embodiments, the liquid crystal and polymer layer can comprise aliquid crystal and polymer network which can be aligned in 3D usingdirectional UV light. In some embodiments the liquid crystal and polymerlayer can be formed at least in part from a Liquid Crystal Polymer (LCP)network. LCPs, which are also referred to in the literature as reactivemesogens, are polymerizable liquid crystals comprising liquidcrystalline monomers containing, for example, reactive acrylate endgroups, which polymerize with one another in the presence ofphoto-initiators and directional UV light to form a rigid network. Themutual polymerization of the ends of the liquid crystal moleculesfreezes their orientation into a three-dimensional pattern. In someembodiments, the process can comprise coating a material systemcontaining liquid crystal polymer onto a substrate and selectivelyaligning the liquid crystal directors using a directionally/spatiallycontrollable UV source prior to annealing. In some embodiments, a liquidcrystal and polymer layer is formed at least in part from a photoaligned layer such as a linearly polymerized photopolymer (LPP). An LPPcan be configured to align LC directors parallel or perpendicular toincident linearly polarized UV light. LPPs can be formed in very thinlayers (typically 50 nm), minimizing the risks of scatter or otherspurious optical effects. Exemplary LPP materials are fabricated byDainippon Ink & Chemicals. In some embodiments, the liquid crystal andpolymer layer can be formed from LCP, LPP and at least one dopant. Aliquid crystal and polymer layer based on LCPs and LPPs can be usedalign LC directors in complex three-dimensional geometries formed in athin film (2-4 microns). In some embodiments, a birefringence controllayer based on LCPs or LPPs further includes dichroic dyes and chiraldopants to achieve narrow or broadband cholesteric filters, twistedretarders, or negative c-plate retarders. Exemplary reactive mesogenmaterials are manufactured by Merck KgaA (Germany). Exemplary LCalignment layer based on LCP are manufactured by Rolic Technologies Ltd.(Allschwil, Switzerland).

Biosensor Architectures Using Raman Signal and Pump Laser Gratings withOrthogonal Grating Vectors

The embodiments described above have Raman signal channels configured aswaveguides aligned parallel to the pump laser channels, with each Ramanchannel containing gratings coupling and conveying to the detector aportion of the spectral bandwidth of the Raman signal excited by oneactive laser pump channel. In other embodiments, the Raman signalgratings can be arranged as a multiplicity of stripes aligned at ninetydegrees to the pump laser channels. The pump laser channels perform thesame function as in the other embodiments. The Raman signal gratingelements are now configured to couple a portion of the spectralbandwidth of the Raman signal excited by one active laser pump channeland focus the Raman signal onto a detector via total internal reflectionwithin a waveguiding substrate.

In one embodiment, shown in FIG. 24 , the laser pump channels in-couplepump laser beams one channel at a time using a laser beam switchingmodule (102), as described above. A SERS layer (106) overlays thesubstrate (154) containing the pump laser channels. The pump lasersubstrate in turn overlays a further substrate (152) containing amultiplicity of grating stripes each having a unique grating vector andgrating pitch for in coupling a specific Raman signal spectral bandwidthscattered from an analyte layer (107) in contact with the SERS layers.Since the grating stripes must diffract long wavelength Raman signal(108), they will tend to have large grating features (typically around5-10 microns for COVID-19 detection applications), in contrast to thepump laser gratings (115) which will have much lower feature spacings(typically sub-micron) for diffracting light (105) in the visible tonear infrared bands. The Raman signal grating elements (153) haveoptical power such that they collimate light the Raman signal (110) inone plane (the waveguide cross-section to enable total internalreflection and focus it onto the detector (112) in the orthogonal plane(the plane of the waveguide)). Hence the grating structure has theprescription of a lens with differing optical power in orthogonal planes(i.e., an anamorphic lens). As in the above described embodiments, thedetector can be a single element device. In some embodiments, thegratings can be arranged so that after in-coupling there are nointeractions with other gratings, to avoid outcoupling. In someembodiments, this can be accomplished by arranging the Raman gratingelements such that they are separated by planar regions which can beused for total internal reflection. In some embodiments, switchinggratings can be used to avoid outcoupling and for eliminating othertypes of unwanted beam/grating interactions.

In some embodiments, as illustrated in FIG. 25 , the pump laser channelsand the Raman signal gratings can be implemented in a single substrate(155). The two types of gratings can be volume or surface reliefgratings, or combinations of the two. In some embodiments, the Ramansignal gratings can be implemented as Fresnel structures. As shown inFIG. 25 , the pump laser grating and the Raman signal gratings can bearranged in alternating stripes on one surface of the substrate. In someembodiments, the two sets of grating stripes can be provided on opposingsides of the substrates. The two gratings having grating vectors inorthogonal directions in the plane of the waveguides, the Raman signalgratings having surface projected grating vectors parallel to the mainpropagation direction of the waveguided signal. Forming the gratings onjust one substrate face is highly desirable in manufacturing terms,since the number of process steps are reduced and the need for gratingalignment is eliminated. In some embodiments, the gratings can betransmission gratings formed on one substrate face. In some embodiments,either or both of the two sets of grating stripes can be configured asreflection gratings. In the case of the Raman signal elements, thereflection gratings can be formed using a molding or stamping processbefore being overcoated by a reflection coating having high reflectivityin the Raman spectral region of interest. The pump beam switching modulecan be similar to the one used in FIG. 24 . A Raman signal elementoperating in transmission can, in some embodiments, be implemented as abinary grating in which the grating feature are printed onto substatesusing a material opaque to the Raman signal wavelengths.

In some embodiments, the SERS coating (106) and the pump laser and Ramansignal gratings (153) can be implemented in a single substrate (155).FIG. 26 shows a portion of a substate supporting pump laser gratingsstripes and Raman signal gratings stripes in which the pump lasergrating stripes are overcoated with a SERS material. In anotherconfiguration shown in FIG. 27 , the SERS overcoated pump laser gratingsand the Raman signal gratings can be disposed on opposing faces of thesubstrate. In some embodiments in which the gratings can be formed usinga low-cost large volume process, the entire substate, including thegratings and the SERS coating, can be treated a s disposable item.

In some embodiments, as an alternative to configuring the pump lasergratings and Raman signal gratings in alternating stripes, the two setsof gratings could be multiplexed. As is apparent from the drawings andthe descriptions of embodiments, the laser and laser beam switchingmodule can be configured in many different ways using various opticalpath folding solutions. In some embodiments, the signal collectionefficiency and signal-to-noise ratio of the detector can be improved byadding at least one of: a detector lens; a light integrator; filters;apertures; and other components commonly used in detection systems.

Embodiments Using Metallic Grating Structures

As discussed above, the present invention can be implemented using manydifferent types of gratings. In some embodiments, gratings based onmetallic structures formed on substrates can offer significantadvantages. One example of such a grating is a wire grid grating. Wiregrid gratings are commonly used as polarizing beamsplitters. In someembodiments, metallic structure gratings can provide either passive orswitching gratings. Metallic structure gratings can provide switchinggratings for use with liquid crystal (LC) and monomer material systemsand other types of electro-optical material systems. Metallic structuregratings can be used to switch volume gratings formed in LC and monomersystems or LC-backfilled surface relief gratings. In some embodiments,metallic structure gratings can be configured to provide in-planeswitching. A further advantage of metallic structure gratings is thatthey can be switched at high frequencies.

Metallic structure gratings can be configured to provide a range ofgrating spatial frequencies to allow operation over the near ultravioletup to LWIR and millimeter wavelength band. Hence, in a biosensor basedon SERS principles, they can provide low spatial frequency gratings forRaman signal collection at wavelengths in the MWIR and LWIR infraredregions and high spatial frequency grating for Raman signal excitationat wavelengths in the UV to near infrared bands. The high refractiveindices of metallic grating structures can be advantageous in manyapplications. In some embodiments, metallic structure gratings can betransmissive or reflective.

In some embodiments, metallic grating structures can be configured toprovide any of the basic grating functionalities described above,including: slanted gratings; rolled-K vector gratings; multiplexedgratings; and chirped gratings. In some embodiments, metallic gratingstructures can be used to apply a thermal stimulus to a layer in contactwith the gratings. In some embodiments, the thermal stimulus can be usedto control refractive index modulation, average index, birefringence, LCalignment and other properties, including their spatio-temporalvariations. In some embodiments, metallic structure gratings can be usedas SERS substrates. In some embodiments, such gratings can be fabricatedin silver or gold.

One exemplary metallic structure grating technology called NanoWeb® ismanufactured by Metamaterials Inc. (Canada). This two-dimensional meshof continuous metal wires can be fabricated onto any glass or plasticsurface. It offers a superior alternative to Indium Tin Oxide (ITO),Silver Nanowire, graphene and carbon nanotube among otherITO-alternative technologies. The design of the mesh geometry allows fora highly conductive and transparent layer. Due to its extremely highconductivity, it is able to operate using very little power whileremaining clear and transparent. Its grid of highly conductive linesallows more energy to pass through an open area surface versusnon-patterned conductive materials. The metal mesh is typically createdfrom silver, aluminum, platinum, copper or nickel. However, almost anytype of metal could be used. The transparency depends only ongeometrical design of the mesh and not the type of metal. Since thelines are of sub-micron thickness, they are effectively invisible to thehuman eye. In some cases, the lines have 500 nm line width with30-micron pitch. Typical specifications are: sheet resistance: from <1to 100 Ohm/sq.; transmission: up to 99%; haze: as low as 1%; line-width:from 0.15 to 1 micron; pitch: 2 microns and above; and thickness: 50 nmto 1 micron. NanoWeb® is manufactured using the Rolling Mask Lithography(RML®) process developed by Metamaterials Inc.

Features of Surface Plasmon Resonance Substrates Used in SomeEmbodiments

Various features of surface plasmons resonance substrates and, inparticular, Surface Enhanced Raman Spectroscopy (SERS) substates can beused in the present invention. SERS substrates are specifically referredto in the following discussion. As discussed above, in some embodiments,such substrates can be formed from nanostructure surfaces ornanoparticles in at least partial contact with layers or assemblies ofdiscrete particles made of noble metals such as silver or gold. In someembodiments, the nanostructures can be three-dimensional photoniccrystal structures formed in LC and monomer material systems afterremoving LC from the cured material. In some embodiments, the photoniccrystals may be recorded in mixtures of monomers and nanoparticles. Thenanoparticles can in some embodiments be made of gold or silver. Thenanoparticles can in some embodiments comprise a nanoparticle componentand Raman reporter material. In some embodiments, the photonic crystalstructures can be Bravais lattices or other regular crystallinestructures. In some embodiments, the photonics crystals may be irregularcrystalline structures.

In some embodiments, the nanostructures can comprise nanocolumns. Insome embodiments, the nanocolumns can have separations of the order of 1micron and feature widths of the order of 10 nanometers. In someembodiments, nanocolumns have high ratios of height to width. In someembodiments, the height of a nanocolumn is of the order of 100 nm. Insome embodiments, the nanostructure can comprise star-shapednanoparticles commonly referred to as nanostars. In some embodiments,stray light from the pump laser beam and fluorescence from the analytecan be blocked using at least one selected from the group of: an opticalfilter; a diffractive optical element; a polarization component; and anoptical absorber. In some embodiments, the fabrication of a SERSsubstrate can include a step of dispersing the analyte and a salt toform agglomerations with average analyte molecule spacings of the orderof 10 nm. In some embodiments, a SERS substate may employ ananostructure, with noble metal applied to the tips of nanostructureelements (for example, nanocolumns or nanostars), that can be deformedby at least one selected from the group of: electrostatic; chemical;mechanical; and electromagnetic means to reduce the spacing of theextremities of nanocolumns, nanostars and other nanostructure featuresto promote high plasmon density.

In some embodiments, the nanostructure can incorporate reportermolecules. In some embodiments, the nanostructure can incorporate alayer of liquid crystal. In some embodiments, the nanostructures canincorporate a nanostructured metallic coating such as a nanoweb or wiregrid grating for use in the application of an electric field that can beused for the modification of average index, index modulation,temperature and other parameters, as discussed above. In someembodiments, the nanostructure features can have spatial, height, aspectratio, cross-section size and separation probability distributions forpromoting plasmon field uniformity or specific hot spot patterns. Insome embodiments, the nanostructure material can incorporate a cappingagent for limiting the growth of the nanofeatures. In some embodiments,nanostructures can be patterned using polystyrene beads. In someembodiments, polystyrene bead can be introduced into a phase separationprocess for forming a nanostructure.

Further Applications of Phase-Separated Diffracting Structures

Embodiments have been described for combining a diffracting structure(typically a nanostructure) with a SERS substrate to provide anintegrated Raman sensor. The phase separation and liquid crystalextraction processes used to make SERS nanostructures can also enable acost-effective solutions for fabricating general-purpose dielectricdiffracting structures using a roll-to-roll process. In someembodiments, phase-separated diffracting structures with feature sizesranging from nanoscales to millimeters can enable many differentapplications extending across the electromagnetic spectrum including theMWIR, LWIR, microwave, and millimeter-wave bands. Since many of thediffracting structures disclosed in below will typically have featuresizes measured in millimeters (for example, structures designed for usewith millimeter waves), the term nanostructure is not generallyapplicable. Therefore, the discussion below will refer to diffractingstructures except in those cases where a nanostructure is the preferredsolution. It should be appreciated that all the diffracting structuresdiscussed below have optical properties based on the electromagnetictheory of gratings. In many embodiments, phase-separated diffractingstructures of any complexity can be synthesized using holographicreplication from masters. In some embodiments, the diffractingstructures are scalable from tiny disposable substrates, such as may beused in the biosensors discussed above, to large area applications invehicles and buildings.

In some embodiments, diverse optical functionalities can be providedusing phase-separated diffracting structures of different opticalprescriptions. For example, there is growing interest in compact devicesthat can integrate sensors and wireless communications. In one suchgroup of embodiments, the present invention can provide a devicecombining phase-separated SERS nanostructures of the type describedabove with phase-separated features of lower spatial frequency that canprovide antennas for wireless communication.

It should be appreciated from the embodiments described that, ingeneral, phase separation can enable a fundamental lithographictechnique that can be applied very cost effectively to very large areasubstrates of any surface geometry. It should also be appreciated fromembodiments to be discussed below that the phase separation processesused to make diffracting structures is not limited to fabricationdiffracting structures, but can also be used to fabricate pixel arraysand associated electronic signal and control circuitry in array devices,with applications in displays and high density liquid crystal displaysin particular, as well as sensors (such as eye tracking, LIDAR and manyothers).

In some embodiments, phase-separated diffracting structures can be usedin combination with diffracting structures fabricated using otherprocesses. In some embodiments, phase-separated diffracting structurescan be combined with diffracting structures with large diffractingfeature sizes suitable for longer wavelength applications. Suchcomposite structures can combine phase-separated diffracting structuresand other diffracting structures on a common surface. In someembodiments, the phase-separated diffracting structures can be combinedwith other types of diffracting structures using multiplexing or inmultiple grating layers.

In some embodiments, a diffracting structure can combine one or morediffracting structures selected from the group of:

-   -   a) a diffracting structure formed by liquid crystal and polymer        phase separation followed by liquid crystal extraction;    -   b) a surface relief diffracting structure fabricated using        nanoimprint lithographic and/or other etching processes;    -   c) a diffracting structure recorded into a holographic        photopolymer material using a holographic exposure process;    -   d) a diffracting structure formed by deposition of a metal or        dielectric layer patterned as a diffracting structure onto a sub        state;    -   e) a transparent metal structure deposited on at least one        substrate for switching a liquid crystal and polymer grating        using transverse fields applied orthogonal to the grating or        fields applied in the plane of the grating (in-plane electric        fields);    -   f) a wavelength diverse diffracting structure;    -   g) a diffracting structure multiplexing at least one of        wavelength and angle;    -   h) a SERS nanostructure;    -   i) a diffracting structure supported by a total internal        reflection waveguide;    -   j) a diffracting structure supported by a substrate transparent        to at least one of the wavelengths diffracted by the diffracting        structures;    -   k) an optical substrate;    -   l) a metal overcoated diffractive structure;    -   m) a dielectric structure incorporating a feature configured as        antenna for providing radiation or detection of free space        propagation or surface propagation of waves;    -   n) a diffracting structure incorporated within a feature        configured as antenna for providing radiation or detection of        free space propagation or surface propagation of waves;    -   o) a diffracting structure operating in transmission; and    -   p) a diffracting structure operating in reflection

Examples of diffracting structures are illustrated in FIGS. 28A-28J andare discussed below. The examples presented should not be construed aslimitations on the scope of the present invention, but rather as anexamples of embodiments thereof. For example, in embodiments wheremultiple layers are specified, the stack order may change. Hence, insome embodiments, the layers illustrated in FIGS. 28A-28J may beinterchanged. In some embodiments, the ray directions shown may bereversed to provide structures that can diffractively in-couple orout-couple electromagnetic radiation. In some embodiments, the gratingstructures can operate in any wavelength band and can be configured foroperation in more than one band. In some embodiments, the opticalstructures illustrated can include additional optical layers forpolarization control, optical filtering, stray light control, opticalpath modification and other functions. In some embodiments, additionallayers including any of the diffractive structures listed above may beprovided. In some embodiments, more than one layer of a given type ofdiffractive structure may be used. In some embodiments, more than onetype of diffracting structure may be formed on a substrate surface.

FIG. 28A illustrates an embodiment comprising a substrate with adiffracting structure formed by depositing a patterned transparent metalor dielectric coating (156) onto an upper surface of the substrate(157). The diffracting structure can diffract an input ray (158)incident within the waveguide into a diffracted ray path (159). In someembodiments, the substrate is a waveguide conveying electromagneticradiation via total internal reflection. In some embodiments, thediffracting structure can diffract input rays transmitted through thelower surface of the substrate.

FIG. 28B illustrates the use of the embodiment of FIG. 28A to diffractan input ray (160) into the waveguide in some embodiments. In someembodiments, the diffracted ray (161) can be coupled into a totalinternal reflection path within the substrate. In some embodiments, thediffracted ray can be transmitted through the lower surface of thesubstrate.

FIG. 28C illustrates an embodiment based on the embodiment of FIG. 28A,further comprising a SERS substrate (106) in contact with a layer ofanalyte (107). As illustrated in FIG. 28C, an input ray (158) which, insome embodiments, is provided by a pump laser beam, can be diffracted(159) out of the substrate towards a point within the SERS/analyteinterface.

FIG. 28D illustrates an embodiment based on the embodiment of FIG. 28C,in which a Raman scatter ray (162) generated within the SERS/analyteinterface is coupled into the substrate as a diffracted ray (163) bymeans of the diffracting structure.

FIG. 28E illustrates an embodiment based on the embodiment of FIG. 28A,in which the substrate contains a switchable diffracting structure (165)formed in a layer. In some embodiments, the switchable diffractingstructure layer can be a liquid crystal and polymer composite. In someembodiments, the substrate can convey total internally reflectedelectromagnetic radiation (167) which can be diffracted (168) out of thesubstrate by the switchable diffracting structure. In some embodiments,an input ray (166) entering the substrate via the lower optical surfacecan be diffracted by the surface grating structure. In some embodiments,the switchable diffracting structure can be based on a non-liquidcrystal based switchable device. In some embodiments, the switchablediffractive structure can employ MEMs technology.

FIG. 28F illustrates an embodiment in which the switchable diffractingstructure layer of FIG. 28E can couple an externally incidentelectromagnetic ray into a total internal reflection path (indicated byrays 170,169) within the substrate.

FIG. 28G illustrates an embodiment based on the embodiment of FIG. 28E,further comprising a SERS substrate in contact with a layer of analyte.As illustrated in FIG. 28G, a ray (168) from a pump laser beam can bediffracted out of the substrate towards a point within the SERS/analyteinterface.

FIG. 28H illustrates an embodiment based on the embodiment of FIG. 28G,in which a Raman scatter ray can be coupled into the substrate by meansof the diffracting structure. In some embodiments, the diffractivestructures formed by depositing a patterned transparent metal ordielectric coating onto an upper surface of the substrate in FIGS.28E-28H can also function as transparent electrodes for switching theswitchable grating structures into diffracting and non-diffractingstates. The electrodes can also be used to provide a continuous range ofdiffraction efficiency over the range from substantially zerodiffraction to a maximum diffraction efficiency. Electrodes formed onone substrate surface, as illustrated in FIGS. 28E-28H, can be used forapplying electric files in directions parallel to the plane of theswitchable grating structure. Such switching schemes are often referredto as in-plane switching. Where it is necessary to apply an electricfield across a switchable grating structure, an additional transparentelectrode can be applied to the lower surface of the substrate. In someembodiments, the lower electrode can be unpatterned. In some embodimentswhere the electric field must have precisely defined directionalcharacteristics, the lower electrode can be patterned. In someembodiments, at least one of the upper or lower electrodes can bediffracting structures designed to provide spatially (in the substrateplane) and directionally varying electrical fields. In some embodiments,electrodes can have electrical and magnetic characteristics that aredynamically reconfigurable using applied electromagnetic fields.

FIG. 28I illustrates an embodiment based on the embodiment of FIG. 28Fin which the upper transparent electrode (171) (which also functions asa diffractive structure for coupling electromagnetic radiation into thesub state) is patterned while the lower transparent electrode (172) isunpatterned. The present invention allows flexibility in the waydiffractive structures can be configured to provide a wide range ofoptical functions. FIG. 28J illustrates an embodiment based on theprinciples discussed above, and is designed to diffract externalelectromagnetic radiation in more than one band while conveying otherelectromagnetic radiation bands via total internal reflection within asubstrate. The apparatus comprises a substrate with an upper transparentelectrode (173) patterned for diffracting first wavelength rays (175)and a lower transparent electrode (174 patterned for diffracting secondwavelength rays (177). A first wavelength diffracted ray (176) is shown.

FIGS. 29A-29F illustrate steps in a process for fabricating aphase-separated partially metallized diffracting structure. FIG. 29Aillustrates an optical substrate (181) coated with a mixture of a liquidcrystal and a monomer (180). FIG. 29B illustrates the exposeddiffracting structure formed after phase separation into polymer-rich(183) and liquid crystal-rich (182) regions has taken place. FIG. 29Cillustrates the diffracting structure (184) formed after evacuation ofliquid crystal. FIG. 29D illustrates a metal coating (185) depositedover the diffracting structure. FIG. 29E illustrates the removal of aportion (186) of the metal coating. FIG. 29F illustrates a planarizedcomposite polymer and metallized polymer diffracting structure (187)formed after removal of the portion of the metal coating. Planarizingthe diffracting structure in this way can facilitate interfacing toother layers such as, for example, SERS substrates and can alsoeliminate non uniformities in roll-to-roll printing processes).

FIG. 30 is a flowchart of steps in a method of fabricating aphase-separated partially metallized diffracting structure, inaccordance with an embodiment of the invention. As shown, the method 190of fabricating a phase-separated partially metallized diffractingstructure is provided. Referring to the flowchart, method 190 includesproviding (191) a substate coated with a mixture of a liquid crystal anda monomer. The mixture can be exposed (192) to form a liquid crystal andpolymer diffracting structure. The liquid crystal component can beevacuated (193) from the diffracting structure. A metal coating can bedeposited (194) on the diffracting structure. A portion of the metalcoat coating can be removed (195) to form a planarize composite polymerand metallized polymer structure.

Millimeter Wave Applications of Phase-Separated Grating Structures

As discussed above, the present invention has many applications in thefield of millimeter wave systems. Examples of millimeter waveapplications using phase-separated grating structures which can employany of the diffractive structures discussed above include:

-   -   a) mirror films for various 5G, 6G applications;    -   b) anti-reflection films (e.g. for increasing transmission into        skin);    -   c) diffractive films with optical power;    -   d) multifunctional films combining gratings for sensing and        wireless communication and other functions and including antenna        electronics real estate combined with SERS nanostructures for        Raman detection;    -   e) frequency agile diffractive structures using switching        gratings.    -   f) angle diversity structures;    -   g) diffractive structures for wireless free-space power        transfer;    -   h) dynamic mirrors;    -   i) synthetic aperture imaging; and    -   j) phased arrays

Gratings Formed Using Coating Processes

In some embodiments, a diffracting structure can be formed using acoating process in which a patterned metal or dielectric layer isapplied to a substrates surface. Examples include:

-   -   a) diffracting structures formed by coating a grating pattern        onto a substrate using masks;    -   b) diffracting structures formed by coating a grating pattern        onto a substrate using an inkjet printing process.    -   c) transparent gratings formed from a conductive material, for        use in either passive or switching modes, deposited on a        substrate;    -   d) diffracting structures that are opaque in one or more        designated bands and transparent in one or more other bands;    -   e) diffracting structures provided by conductive material        deposited onto a substrates for the purposes of switching a        layer containing liquid crystal or switching a grating formed        from a liquid crystal and monomer material system;    -   f) electrodes that can be used for switching polymer/LC        diffracting structures whilst providing an opaque grating        structure for diffracting Raman scattered light from an        analyte/SERS interface or diffracting the pump laser light into        the analyte/SERS interface;    -   g) electrodes as described above configured to provide either        orthogonal or in-plane electric fields

FIG. 31 illustrates a biosensor based on the embodiment of FIG. 25 . TheRaman signal grating elements and pump laser gratings are replaced inFIG. 31 by metal or dielectric grating elements (198) formed used acoating process on the substrate 197. The surface diffraction structurescan provide electrodes for switching a liquid crystal and polymerdiffracting formed within the waveguide as discussed above. In someembodiments, the substrate (197) can incorporate a switchablediffractive structure (196).

Reconfigurable Diffracting Structures

Due to their large birefringence and moderately low loss, liquidcrystals (LCs) are a promising dielectric media for development of avariety of reconfigurable and tunable devices extending across theelectromagnetic spectrum including infrared microwaves andmillimeter-wave with properties that can be designed by suitable choiceof dielectric and elastic constants and other LC parameters. Devicescombining LC material with diffractive structures offer potential fordevices such as angle and frequency selective, adaptive arrays, beamsteering and many others.

Liquid crystals possess the smallest elastic constants and the largestbirefringence, spanning the visible to infrared bands and beyond, amongall known materials. Liquid crystals can be chemically synthesized andprocessed on a very large scale, and they are also compatible withalmost all technologically important optoelectronic materials. Liquidcrystals can be controlled by electricity, light, acoustic waves, andother means. Efficient and versatile drive and switching techniques havebeen developed over the years. By integrating liquid crystals withplasmonic nanostructures, active plasmonic materials and devices withenhanced performance have been demonstrated. As discussed above, liquidcrystal can be combined with polymers to enable high index modulationswitching, spatio-temporally varying refractive index modulation andversatile means for integrating complex diffractive optical devices,including Bragg gratings. In some embodiments, the liquid crystalpolymer devices can be augmented by nanoparticles to provide theelectrooptical properties.

Phase Separation as a Large Area Lithographic Process

Phase-separated diffracting structures can provide high resolutionswitchable arrays for use in displays including liquid crystal displaysand other types of spatial light modulators. Light field displays, forexample, require high pixel density arrays with pitches as low as 1-2microns to achieve acceptable field of view, image resolution andeyebox. Arrays with such feature resolutions are not cost effective tomanufacture using current volume processes for depositing transparentelectrodes (using materials such as ITO). Roll-to-roll lithographicprocesses, such as RML, suffer from periodic pitch variations due to thecompressibility of rollers making lining up array elements with drivecircuitry very challenging. FIG. 32 illustrates elements of an activematrix thin film transistor (200) type array in which the pixelelectrodes (199) and signal (202) and control circuitry (201) are formedusing a phase separation process similar to the one used for fabricatingthe phase-separated diffracting structures discussed above. The arrayscan be of any size and can be formed on curved substrates. The requiredarray pixel sizes, pitches and circuitry dimensions required in highdensity light field displays and in many other array applications arewell within the resolution capability of the phase separation technique.

Phase Separation Structures for Wireless Applications

Wireless communication applications require an antenna to enablefunctionalities such as beam steering, direction finding, radar, etc.Reconfigurable antennas can switch between functions using a singlestructure eliminating the need for multiple antennas. Antennareconfiguration to provide different functionalities can be achievedthrough a change in the antenna's geometry and/or electrical behavior.Reconfigurable antennas typically have two or more discretely orcontinuously switchable states. These different states are normallyobtained by changing current paths of the antenna through eitherrearranging the antenna itself or altering its surrounding medium.Reconfigurable antennas have been applied in radiofrequency (RF) systemsfor wireless and satellite communication, imaging and sensing. In someembodiments, a reconfigurable antenna can be provided by incorporating adynamically reconfigurable diffracting structure including a liquidcrystal component.

FIG. 33 conceptually illustrates one example of an antenna (204) whichincorporates a reconfigurable diffracting structure, including a liquidcrystal component (203). In such embodiments, liquid crystal diffractingstructures can be used to modify local currents of a metal-based antennato reconfigure its radiation parameters.

In some embodiments, nanostructures for use with SERS substrates can beformed from a phase separation process starting from a mixture of liquidcrystal, monomer and nanoparticles (typically silver or gold). Aftercuring has been completed, the liquid crystal can be extracted to leavenanostructure comprised polymer regions within which nanoparticles aredispersed. Various grating structures, which are essentially differentforms of photonic crystal, can be formed as illustrated in FIGS. 34-36 .

FIG. 34 shows a simple phase-separated Bragg grating structure (205)comprising polymer fringes (207) immersed in air (208) with each fringecontaining nanoparticles (208) dispersed within a polymer matrix (209).FIG. 35 shows a slice through a three-dimensional phase-separatedphotonic crystal or lattice (210) comprising polymer regions (211)immersed in air (212), with each polymer region containing nanoparticles(213) dispersed within a polymer matrix (214). FIG. 36 shows aphase-separated multiplexed grating structure (215) from first (216) andsecond (217) crossed Bragg gratings immersed in air (220), where eachgrating comprises fringes containing nanoparticles (218) dispersedwithin a polymer matrix (219). Many other nanostructure configurationscan be provided based on the description and figures provided.

The diffractive structures illustrated in FIGS. 34-36 may have morecomplex morphologies than the ones illustrated. In some embodiments, theevacuated regions resulting from the evacuation of liquid crystal maycontain a residual polymer network, including gold or silvernanoparticles suspended in the polymer network. FIG. 37 illustrates sucha phase-separated diffractive structure based on the embodiment of FIG.36 . In the embodiment of FIG. 37 , the evacuated regions now comprisethe residual polymer network (222) containing dispersed nanoparticles(223). The concentration of polymer in the evacuated regions willtypically be much lower than that of the polymer-rich regions. In someembodiments, the relative concentration of nanoparticles in thepolymer-rich regions and the intervening regions may differ. In someembodiments, the nanoparticles may be uniformly dispersed in the film,while the polymer grating may be periodic. There are no restrictions onthe type of polymer grating that can be formed. Any of the grating formsdiscussed above may be used in various embodiments based on theembodiments illustrated in FIGS. 34-36 . In some embodiments, thegrating prescription and concentration of nanoparticle concentration canhave spatial variations that are not limited to uniform or periodic. Insome embodiments of a biosensor for saliva testing, the analyte (saliva)may only penetrate the low concentration polymer and nanoparticleregions. Such embodiments offer the dual benefits of a diffractivestructure that can be configured for various purposes, such as routingof light, light concentration etc., and a uniform dispersion ofnanoparticles that act as surface plasmon resonance surfaces foramplifying the Raman scatter from the analyte. In some embodiments, thenanoparticles may incorporate reporter molecules as discussed above. Thenanoparticles are not limited to any particular size or shape. In someembodiments, the nanoparticles may comprise particles of more than onedifferent size or shape.

In some embodiments, it may be advantageous to form nanostructures ofthe type disclosed from polymers. However, it is known that somepolymers can exhibit fluorescence under exposure to visible light. Thefluorescence can reduce signal to noise ratio in Raman spectroscopy. Theeffect can be reduced by using high functionality monomers in thefabrication of the grating. In some embodiments, the biosensor will usepump lasers emitting in the blue region. However, in some embodimentsgratings can be fabricated using high functionality monomer materialsystems designed for broader range of visible wavelengths or forinfrared wavelengths.

The foregoing embodiments and advantages are merely exemplary, and arenot to be construed as limiting the present invention. The presentteaching can be readily applied to other types of apparatuses. Thedescription of the present invention is intended to be illustrative, andnot to limit the scope of the claims. Many alternatives, modifications,and variations will be apparent to those skilled in the art. Variouschanges may be made without departing from the spirit and scope of theinvention, as defined in the following claims.

What is claimed is:
 1. A molecular sensor, comprising: a pump lasersource that emits pump light having at least one wavelength; a surfaceenhanced Raman spectroscopy (SERS) substrate; at least one type ofmolecule disposed in proximity to said SERS substrate, each of said atleast one type of molecule exhibiting a unique Raman spectrum underirradiation from said light at said at least one wavelength; a detectorwith a detection bandwidth covering a Raman spectrum of said molecule; apump laser substrate comprising at least one pump laser channel forpropagating pump laser beams; a pump beam switch for directing a portionof the pump light from said pump laser source into each of said pumplaser channels sequentially; a coupling layer overlaying each of said atleast one pump laser channel for directing the pump light portionpropagating in each channel towards said SERS substrate; and a Ramansignal detection substrate comprising at least one Raman signaldetection channel supporting an optical structure for selectivelycoupling in and transmitting towards said detector a portion of a Ramanspectrum emitted by said at least one type of molecule after excitationby said portion of the pump light.
 2. The molecular sensor of claim 1,wherein said optical structure is formed on at least one optical surfaceof said Raman signal detection substrate.
 3. The molecular sensor ofclaim 1, wherein the Raman signal detection substrate comprisesreflective optical surfaces formed on at least one of the upper andlower surfaces of said Raman signal substrate, and wherein thereflective optical surfaces include optical structures configured fordirecting said Raman signal along said Raman signal detection substratetowards the detector using at least one internal reflection.
 4. Themolecular sensor of claim 1, further comprising a Raman signal combinerfor coupling each of said at least one Raman signal detection channel tosaid detector.
 5. The molecular sensor of claim 1, further comprising ananostructured substrate optically coupled to said SERS substrate,wherein said nanostructured substrate comprises at least one selectedfrom the group of: a substrate supporting nanoparticles; a substratesupporting a nanostructure with a spatially varying nanostructurefrequency; and a substrate supporting a nanostructure with a spatiallyvarying nanostructure amplitude.
 6. The molecular sensor of claim 1,wherein the portion of said coupling layer overlaying each said pumplaser channel has a prescription for satisfying a Raman scatteringmomentum balance in a SERS surface plasmon region for one type ofmolecule.
 7. The molecular sensor of claim 1, wherein at least a portionof said Raman signal detection substrate has optical power.
 8. Themolecular sensor of claim 1, wherein said coupling layer comprisesnanostructures configured to provide surface plasmon characteristics forstimulating and amplifying Raman scattering from said at least onemolecule.
 9. The molecular sensor of claim 1, wherein: (1) said couplinglayer and said Raman signal detection substrate are combined in a commonsubstrate; (2) said common substrate includes a diffracting structureconfigured for directing the pump light portion propagating in eachchannel towards said SERS substrate; (3) said common substrate includesan optical structure formed on at least one optical surface of saidcommon substrate for selectively coupling in a portion of a Ramanspectrum emitted by said at least one type of molecule after excitationby said portion of the pump light; and (4) said common substrate furthercomprises reflective optical surfaces formed on at least one of theupper and lower surfaces of said common substrate, and the reflectiveoptical surfaces include optical structures configured for directingsaid Raman signal along said common substrate towards said detectorusing at least one internal reflection.
 10. The molecular sensor ofclaim 9, wherein said nanostructure substrate and said opticalstructures are aligned in orthogonal directions.
 11. The molecularsensor of claim 1, wherein said pump beam switch comprises at least oneswitching grating.
 12. The molecular sensor of claim 9, wherein saidnanostructure substrate is configured for optimizing surface plasmonswithin regions of said SERS substrate in proximity to said at least onetype of molecule.
 13. The molecular sensor of claim 1, wherein said SERSsubstrate incorporates at least one selected from the group of: areporter molecule; nanostructures structures formed using a phaseseparation process; metallized nanostructures structures; and more thanone type of diffracting structure.
 14. The molecular sensor of claim 1,wherein an output signal from said detector is coupled to a smartphonefor processing and display of Raman spectra.
 15. The molecular sensor ofclaim 1, configured as compact Raman spectrometer.
 16. The molecularsensor of claim 1, further comprising nanostructures for providingreconfigurable diffractive antennas for at least one selected from thegroup of: wireless communications; long wavelength electromagneticradiation collection; and detector coupling.
 17. The molecular sensor ofclaim 1, configured for detecting COVID-19 from saliva using at leastone of multivariate analysis of selected Raman spectrum lines ormeasurement of a Dublin-Boston score.
 18. The molecular sensor of claim1, further comprising grating structures operating in the millimeterwave band.
 19. The molecular sensor of claim 1, further comprising agrating structure formed from a high functionality monomer andexhibiting a low fluorescence cross-section when irradiated by said pumplight at said at least one wavelength.
 20. The molecular sensor of claim1, further comprising at least one selected from the group of: a liquidcrystal layer disposed in proximity to the SERS substrate; at least onemicrofluidic component; magnetic components for manipulation of said atleast one type of molecule; nanoparticles for manipulation of said atleast one type of molecule; and a pump laser source emitting blue light.